ST2⁺ Tregs preferentially home outside of secondary lymphoid organs and exhibit a highly activated phenotype.

Abstract

<p>Flow cytometric analysis of the phenotype and frequency of WT ST2⁺ and ST2⁻ Foxp3⁺ Tregs in spleen, pLN, blood, lung, lamina propria of the small intestine (siLP) and colon (coLP): <i>(A)</i> Frequency of ST2⁺ Tregs <i>(left)</i> and MFI of the ST2 staining on the ST2⁺ Treg fraction <i>(right)</i>. <i>(B)</i> MFI of chemokine receptor and α4β7 staining on ST2⁺ and ST2⁻ Tregs. <i>(C)</i> KLRG1 and CD103 expression in ST2⁺ <i>(top)</i> and ST2⁻ <i>(bottom)</i> Tregs from spleen; quantified frequencies from indicated organs <i>(right)</i>. <i>(D)</i> Frequency of CD44^hi, CD62L^lo and CTLA-4⁺ T cells within ST2⁺ and ST2⁻ Treg populations. <i>(E)</i> MFI of the Foxp3 staining <i>(left)</i> and geometric mean index of GATA-3 <i>(right)</i> in ST2⁺ and ST2⁻ Tregs. <i>(F)</i> Quantification of mRNA expression of the indicated genes from FACS-sorted ST2⁺ and ST2⁻ CD25⁺ Tregs from spleen and pLN <i>ex vivo</i>. mRNA expression normalized to <i>Hprt</i> endogenous control. <i>(G)</i> Frequency of ST2⁺ and ST2⁻ Tregs with IL-10 production capability as detected by GFP expression from <i>B6</i>.<i>Foxp3</i>^<i>hCD2</i> <i>xIl10</i>^<i>gfp</i> reporter mice. Fig <i>2A</i>: Data are representative of at least 2 independent experiments. Bar graphs show the mean ± SD of at least 5 biological replicates. Fig <i>2B</i>: pooled data from 2 independent experiments with 3–5 biological replicates each. Bar graphs show the mean ± SD. Fig <i>2C–2E</i> and <i>2G</i>: Data are representative of at least 2 independent experiments. Scatter plots depict one mouse as individual dot with mean ± SD. Fig <i>2F</i>: pooled data from 2 independent experiments. Significance was tested using unpaired Student’s t test. * p ≤ 0.05; ** p ≤ 0.01; *** p ≤ 0.001; non-significant (ns) p > 0.05.</p